DESCRIPTION: (Provided by the applicant): Our ability to control mosquitoes that transmit human diseases is threatened by pesticide resistance. One common form of pesticide resistance, metabolic resistance, is frequently associated with the over-expression of detoxification enzymes. Numerous detoxification enzyme genes have been cloned from resistant insects, but rarely do these genes map to a resistance locus. Evidence strongly suggests that one of the major resistance loci in insects is a yet undefined trans-regulatory gene that controls the expression of detoxification enzymes, such as cytochrome P450s. The long-range goal of our laboratory is to understand the molecular basis of metabolic pesticide resistance in vector insects, such as mosquitoes. A well-established approach for identifying resistance genes in pest insects is to first discover the genes in Drosophila. Since mosquitoes and Drosophila share a high degree of homology across their genomes, and the Drosophila genome is now sequenced, the quickest path to identifying a gene of key importance in metabolic resistance in mosquitoes is to first characterize the gene in Drosophila. The major metabolic resistance locus in Drosophila is a dominant trait known as Rst (2) DDT. This locus has been mapped to a region encompassing 35 previously sequenced genes, but to date we do not know which gene actually confers the metabolic resistance phenotype. The objective of the work proposed here is to identify a gene involved in metabolic resistance in mosquitoes by first determining its orthologue in Drosophila. The central hypothesis of this grant is that metabolic resistance to insecticides in Drosophila is due to one of these 35 known genes. This hypothesis is based on recombinational mapping data showing that this narrow region of the second chromosome is consistently associated with metabolic resistance to pesticides. The three specific aims of this project are as follows. First, identify candidate resistance genes in Drosophila by determining open reading frames that have structural or expression level differences between the resistant and susceptible insects. Second, establish which of the candidate genes is/are causally responsible for metabolic resistance by transforming the resistant allele into a susceptible Drosophila line. To determine which gene confers resistance, pesticide bioassays will be performed on the transformed flies. Only those insects transformed with the gene for resistance will survive the bioassay. Third, identify the orthologue of this gene in Anopheles gambiae. We expect to identify a gene that plays a crucial role in metabolic resistance in Dipterans. The proposed research is significant because this gene will make an excellent target-site for the discovery of compounds capable of suppressing metabolic resistance in vector species that transmit human diseases.